BACKGROUND AND AIMS: The red material occupying the larger portion of the acquired sample in EUS fine needle biopsy (FNB) is seldom investigated. We aimed to evaluate the composition of the red material. METHODS: Patients with a solid pancreatic mass who received EUS FNB from Sep. 2020 to Jun. 2021 were enrolled. The white and red materials were separated immediately after puncture. Needle passes proceeded until two specimens with more than 4 mm of white material were obtained. An extra needle pass was conducted for DNA collection. The DNA amount, KRAS mutation type and mutation allele frequency were compared between the white and red material. RESULTS: A total of 40 patients were enrolled with 68 paired white and red materials. The diagnostic accuracy was slightly higher in the white material (92.5% vs. 82.5%, p=0.219). On the histology slides, the area of the tumor gland was comparable in both materials, but the total tissue area was larger in the red material (9.74 & 10.74 mm(2) larger by GLM &GEE, both p<0.001). The amount of DNA was significantly higher in the red material (2.99 μg [IQR 1.67-7.17] vs. 0.70 μg [IQR 0.30-1.19], p<0.001). Common pancreatic adenocarcinoma KRAS mutation was identified at a rate of 85% for the white material and 95% for the red material. Whether it was red or white material, there was a high concordance of KRAS mutation types (34/40, 85%) and high correlation of mutation allele frequency (ρ=0.66, p<0.001). CONCLUSIONS: In EUS FNB, the red material contains a higher amount of tumor DNA, and can be an alternative source for tumor DNA analysis.